DESCRIPTION: (Applicant's Abstract) Drug addiction is a major problem in the developed world. It affects mostly young people and destroys lives. To understand the reinforcing and motivational aspects of drug abuse, we need to elucidate the biological and molecular mechanisms that are at work in drug addiction. Cocaine and amphetamine exert their addictive properties via the dopaminergic system. Repeated abuse of these drugs causes the long-term behavioral consequences of drug addiction, drug tolerance, and drug sensitization. These adaptations develop over time, persist long after the cessation of drug use, and involve long-term changes in dopaminoceptive neurons. The biological substrate of these changes must be searched in the effect these drugs have on gene expression. Since cocaine and amphetamine work in a D1 dopamine receptor-dependent manner, the investigation of the second messenger pathway from D1 receptor stimulation to gene expression can provide us with insights into the mechanism of drug addiction. The goal of the present proposal is to elucidate how the signal at the D1 receptor at the synapse directs gene regulation in the nucleus. We hypothesize that dopamine activates a combination of factors that cause depolarization and that, altogether with second messengers inside the neuron, effect gene expression. We have characterized a model system of gene expression in primary striatal cultures that is activated by drug abuse. With the help of this system we have identified elements of the signaling cascade. An important contributor to dopamine signaling identified in our lab is the N-methyl-D-aspartate (NMDA) receptor. D1 receptor-mediated gene expression is blocked by NMDA antagonists, and is dependent on functional NMDA receptors. It is not clear how both receptors are coupled. Preliminary data that show that dopamine causes phosphorylation of the NR1 subtype of the NMDA receptor provide a potential mechanism of activation. We now propose to further characterize the role NR1 phosphorylation may play in D1 receptor-mediated gene expression. Moreover, we will investigate how the NMDA receptor transduces the dopamine signal to the neuronal nucleus. Analysis of NMDA receptor-mediated gene expression points to an involvement of L-type Ca2+ channels. Since D1 receptor-mediated gene expression depends on NMDA receptors, we will examine the role of L-type Ca2+ channels in D1 receptor-mediated gene expression. The role of intracellular kinases and phosphatases in D1 receptor-mediated gene expression will also be addressed. The elucidation of the D1 receptor-mediated signaling cascade is crucial to advance our understanding of the mechanism of drug addiction. In addition, valuable insight into novel therapeutic strategies to prevent or reverse addiction may be gained.